Anaerobic regulation of the Escherichia coli dmsABC operon requires the molybdate-responsive regulator ModE.
Expression of the Escherichia coli dmsABC operon that encodes a molybdenum-containing DMSO/TMAO reductase is increased in response to anaerobiosis and repressed by nitrate. These changes are mediated by the transcription factors Fnr and NarL respectively. Interestingly, modC strains that are defective in molybdate uptake exhibit impaired anaerobic induction and no nitrate-dependent repression of the dmsABC operon. To determine if the molybdate-responsive transcription factor ModE is involved in this process, a set of dmsA-lacZ operon fusions were constructed and analysed. The pattern of dmsA-lacZ expression in response to anaerobiosis and nitrate addition was identical in both modC and modE strains, thus suggesting a regulatory role for ModE. In vitro studies confirmed that ModE bound the dmsA promoter at a high-affinity site typical of other E. coli ModE operator sites. Mutations in this site abolished ModE binding in vitro and displayed the same phenotype as a modE mutation. In contrast to previously characterized ModE operator sites, which either overlap or are located immediately upstream of the ModE-regulated promoter, the ModE site is centred 52.5 bp downstream of the major dmsA transcript start site. We identified a putative integration host factor (IHF) binding site in the intervening sequence, and in vitro studies confirmed that IHF bound this site with high affinity. Using himA mutants, we confirmed that IHF plays a role in the molybdate-dependent regulation of dmsA-lacZ expression in vivo. This study provides the first example in which ModE affects gene regulation in concert with another transcription factor.
McNicholas
P M
PM
Department of Microbiology and Molecular Genetics and the Molecular Biology Institute, University of California at Los Angeles, 90095-1489, USA.
Chiang
R C
RC
Gunsalus
R P
RP
D000693:Anaerobiosis; D001426:Bacterial Proteins / Q000737:chemistry; D001483:Base Sequence; D018983:DNA Footprinting; D004269:DNA, Bacterial / Q000737:chemistry; D004268:DNA-Binding Proteins / Q000737:chemistry; D003850:Deoxyribonuclease I; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins; D005696:Galactosidases / Q000032:analysis; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology*; D034982:Integration Host Factors; D008969:Molecular Sequence Data; D008982:Molybdenum / Q000737:chemistry*; D009566:Nitrates / Q000737:chemistry; D009876:Operon / Q000502:physiology; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D014157:Transcription Factors / Q000737:chemistry / Q000235:genetics / Q000502:physiology*
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D034982:Integration Host Factors; C491064:ModE protein, E coli; C101050:ModE protein, bacteria; D009566:Nitrates; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; C044659:molybdate; D008982:Molybdenum; D003850:Deoxyribonuclease I; D005696:Galactosidases